Bio-rad Aurum™ Total RNA Mini Kit Manual do Utilizador

Aurum™Total RNA Mini Kit Instruction ManualCatalog # 732-6820For technical support, call your local Bio-Rad office, orin the US, call 1-800-4BIORAD (1

6Elution Guidelines• Apply elution solution directly to the membrane stack at the base of each RNAbinding columnRibonucleases• Although the components

7Table 2. Disruption and homogenization methods.Starting Disruption Homogenization Material Method MethodCultured mammalian Lysis solution Pipetti

8Section 6Vacuum Manifold Setup and Use With the Column Adaptor Plate (CAP)Guidelines for Vacuum Format • The recommended operating range is –17 to –2

9Preparing the Aurum Vacuum ManifoldTubing provided in the Aurum vacuum manifold kit (catalog # 732-6470) is 4 ftlong and must be cut into appropriate

10Manifold Wash Setup (Figure 2)1. Insert the CAP (luer ends up) into the depression in the vacuum manifoldtop. Ensure that the CAP rests evenly on t

Section 7Vacuum ProtocolImportant: Please read Section 5, “Before Using the Aurum™ Total RNAMini Kit” and Section 6 "Vacuum Manifold Setup and Us

12Bacteria Follow steps B1–B4, then continue with step 1 of “All Starting Sample Types”on page 13.B1. Transfer up to the equivalent of 3 OD•ml bacter

13Animal and Plant TissueFollow steps D1–D5, then continue with step 1 of "All Starting Sample Types"on page 13.D1. Cut the tissue into smal

144. Add 700 µl of low stringency wash solution to the RNA binding columnand close the vacuum regulator dial until the gauge indicates –17 to –23 inHg

15Section 8Spin ProtocolImportant: Please read Section 5, “Before Using the Aurum™ Total RNA MiniKit,” before proceeding.The Aurum total RNA mini kit

4110133B:4110133A.qxd 1/16/2009 2:35 PM Page TOCi

B3. Add 350 µl of lysis solution (already supplemented with 1% b-mercapto-ethanol) to each tube. Pipet up and down several timesto mix thoroughly.B4.

17D2. Transfer up to 20 mg hard animal tissue, up to 40 mg soft animal tissue,or up to 60 mg plant tissue into an RNase-free 2.0 ml cappedmicrocentrif

186. For each column processed, mix 5 µl of reconstituted DNase I with 75 µlof DNase dilution solution in a 1.5 ml microcentrifuge tube (not provided)

Section 9Troubleshooting GuideProblem Possible Cause Recommended SolutionGenomic DNA Incomplete DNase I Increase DNase I digestcontamination digest

20Problem Possible Cause Recommended SolutionLow eluate volume Insufficient centrifugation Add 1–3 min to the(<60 µl) time during elution centrif

Problem Possible Cause Recommended SolutionTotal RNA prep Incorrect use of wash Add the appropriate performs poorly solutions volume of 95–100% in dow

Section 10Ordering InformationCatalog # Description732-6820 Aurum Total RNA Mini Kit732-6830 Aurum Total RNA Fatty and Fibrous Tissue Kit732-6870 Auru

4110133B:4110133A.qxd 1/16/2009 2:35 PM Page 23

Bio-Rad Laboratories, Inc.2000 Alfred Nobel Dr.Hercules, CA 94547 USA(510) 741-10001-800-424-67234110133 Rev BLife ScienceGroup00-0000 0000

Table of ContentsSection 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . .1Section 2 Kit Components . . . . . . . . . . . . .

4110133B:4110133A.qxd 1/16/2009 2:35 PM Page TOC2

Section 1Introduction The Aurum™ total RNA mini kit purifies total RNA samples rapidly frommammalian cell cultures, bacteria, and yeast (Saccharomyces

Section 3Storage ConditionsAll kit components (including lyophilized DNase I) should be stored at roomtemperature. Store reconstituted DNase I at –20°

Section 5Before Using the Aurum Total RNA Mini Kit Please read the following guidelines before proceeding with the total RNApurification.Starting Mate

Table 1. Yield (per column) of total RNA from various samplesusing the Aurum total RNA mini kit. Starting Material Avg. Yield (µg)*Cultured cells (2 x

5Reagents Used With the Aurum Total RNA Mini Kit• The low stringency wash solution is provided as a 5x concentrate. Add 4 volumes (80 ml) of 95–100% e